Bottom Line:
One such satratoxin chemotype-specific cluster is adjacent to the core trichothecene cluster, which has diverged from those of other trichothecene producers to contain a unique polyketide synthase.The results suggest that chemotype-specific gene clusters are likely the genetic basis for the mutually-exclusive toxin chemotypes of Stachybotrys.A unified biochemical model for Stachybotrys toxin production is presented.

Background: The fungal genus Stachybotrys produces several diverse toxins that affect human health. Its strains comprise two mutually-exclusive toxin chemotypes, one producing satratoxins, which are a subclass of trichothecenes, and the other producing the less-toxic atranones. To determine the genetic basis for chemotype-specific differences in toxin production, the genomes of four Stachybotrys strains were sequenced and assembled de novo. Two of these strains produce atranones and two produce satratoxins.

Results: Comparative analysis of these four 35-Mbp genomes revealed several chemotype-specific gene clusters that are predicted to make secondary metabolites. The largest, which was named the core atranone cluster, encodes 14 proteins that may suffice to produce all observed atranone compounds via reactions that include an unusual Baeyer-Villiger oxidation. Satratoxins are suggested to be made by products of multiple gene clusters that encode 21 proteins in all, including polyketide synthases, acetyltransferases, and other enzymes expected to modify the trichothecene skeleton. One such satratoxin chemotype-specific cluster is adjacent to the core trichothecene cluster, which has diverged from those of other trichothecene producers to contain a unique polyketide synthase.

Conclusions: The results suggest that chemotype-specific gene clusters are likely the genetic basis for the mutually-exclusive toxin chemotypes of Stachybotrys. A unified biochemical model for Stachybotrys toxin production is presented. Overall, the four genomes described here will be useful for ongoing studies of this mold's diverse toxicity mechanisms.

Fig5: The core atranone clusters of theStachybotrysatranone-producing strains. The core atranone clusters are shown in the blue box. The other genes shown are chemotype-independent. ATR12 of strain 40288 is gray to indicate that it is a possible pseudogene, because despite its translation having ~90% identity to 40285 Atr12, in the present assembly its exon 1 contains an internal stop codon.

Mentions:
The products of the core atranone cluster likely suffice to make all known atranone species. The hypothesis of this study was that the two mutually-exclusive chemotypes of Stachybotrys were due to the presence of strain-specific SMB clusters. To test this hypothesis computationally, the four Stachybotrys genome assemblies were searched for loci that were present in both satratoxin strains but in neither atranone strain, or vice versa. The custom search strategy combined two methods, both based on sequence alignment. At the genomic level, four-way whole-genome alignment was employed, using Mugsy [32]. At the level of the proteome, the sets of homologs compiled with OrthoMCL were considered. Whole-genome alignment was needed to show genomic context, but in practice Mugsy aligned some locus boundaries incorrectly, so its results were manually adjusted as described in the Methods. Overall, the search yielded a total of two atranone-specific and four satratoxin chemotype-specific gene clusters. The larger of the two atranone-specific gene clusters was named the core atranone cluster (CAC, or AC1; Figure 5, Additional file 5). This is a ~35-kbp PKS-based cluster, and it has a nearly-identical architecture of 13–14 genes (ATR1-ATR14) in both atranone strains. The CAC is complete in the sense that the genes immediately flanking it on both sides are not atranone-specific.Figure 5

Fig5: The core atranone clusters of theStachybotrysatranone-producing strains. The core atranone clusters are shown in the blue box. The other genes shown are chemotype-independent. ATR12 of strain 40288 is gray to indicate that it is a possible pseudogene, because despite its translation having ~90% identity to 40285 Atr12, in the present assembly its exon 1 contains an internal stop codon.

Mentions:
The products of the core atranone cluster likely suffice to make all known atranone species. The hypothesis of this study was that the two mutually-exclusive chemotypes of Stachybotrys were due to the presence of strain-specific SMB clusters. To test this hypothesis computationally, the four Stachybotrys genome assemblies were searched for loci that were present in both satratoxin strains but in neither atranone strain, or vice versa. The custom search strategy combined two methods, both based on sequence alignment. At the genomic level, four-way whole-genome alignment was employed, using Mugsy [32]. At the level of the proteome, the sets of homologs compiled with OrthoMCL were considered. Whole-genome alignment was needed to show genomic context, but in practice Mugsy aligned some locus boundaries incorrectly, so its results were manually adjusted as described in the Methods. Overall, the search yielded a total of two atranone-specific and four satratoxin chemotype-specific gene clusters. The larger of the two atranone-specific gene clusters was named the core atranone cluster (CAC, or AC1; Figure 5, Additional file 5). This is a ~35-kbp PKS-based cluster, and it has a nearly-identical architecture of 13–14 genes (ATR1-ATR14) in both atranone strains. The CAC is complete in the sense that the genes immediately flanking it on both sides are not atranone-specific.Figure 5

Bottom Line:
One such satratoxin chemotype-specific cluster is adjacent to the core trichothecene cluster, which has diverged from those of other trichothecene producers to contain a unique polyketide synthase.The results suggest that chemotype-specific gene clusters are likely the genetic basis for the mutually-exclusive toxin chemotypes of Stachybotrys.A unified biochemical model for Stachybotrys toxin production is presented.

Background: The fungal genus Stachybotrys produces several diverse toxins that affect human health. Its strains comprise two mutually-exclusive toxin chemotypes, one producing satratoxins, which are a subclass of trichothecenes, and the other producing the less-toxic atranones. To determine the genetic basis for chemotype-specific differences in toxin production, the genomes of four Stachybotrys strains were sequenced and assembled de novo. Two of these strains produce atranones and two produce satratoxins.

Results: Comparative analysis of these four 35-Mbp genomes revealed several chemotype-specific gene clusters that are predicted to make secondary metabolites. The largest, which was named the core atranone cluster, encodes 14 proteins that may suffice to produce all observed atranone compounds via reactions that include an unusual Baeyer-Villiger oxidation. Satratoxins are suggested to be made by products of multiple gene clusters that encode 21 proteins in all, including polyketide synthases, acetyltransferases, and other enzymes expected to modify the trichothecene skeleton. One such satratoxin chemotype-specific cluster is adjacent to the core trichothecene cluster, which has diverged from those of other trichothecene producers to contain a unique polyketide synthase.

Conclusions: The results suggest that chemotype-specific gene clusters are likely the genetic basis for the mutually-exclusive toxin chemotypes of Stachybotrys. A unified biochemical model for Stachybotrys toxin production is presented. Overall, the four genomes described here will be useful for ongoing studies of this mold's diverse toxicity mechanisms.